Previous therapeutic approaches to prevent sickle red cell dehydration have targetted KCI cotranspot and K and CI conductances. I propose to develop the non-selective cation pathway as a therapeutic target. I have recorded unitary openings and closings of this pathway in sickle red cells and have found it to be activated by deoxygenation. I propose here to characterize the biophysical and pharmacologic properties of the channel. I will characterize the mechanisms of its activation by dehydration and will attempt to define the role of HbS polymerization in that activation. We will also define other possible modes of regulation by cell volume, by redox regulation and by small molecule or protein signalling pathways. We will compare this channel of human sickle cells to hypothesized counterparts in red cells from a mouse model of sickle cell disease. The information gained from these studies will assist in the molecular identification of the channel polypeptide and will accelerate development of this channel as a therapeutic target. The specific aims of the proposal are: (1) Test the hypothesis that the erythroid cation channel is uprequlated or dysrequlated in sickle cell disease. The experiments of this aim will define the biophysical and pharmacological properties of cation channel activity in HbSS and HbAA erythrocytes. They will further define the stimulatory and inhibitory regulators of these channel activities, with a particular focus on hypoxia as a stimulatory regulator. (2) Test the hypothesis that stimulation of cation channel activity in HbSS erythrocytes by deoxyqenation is mediated by or requires polymerization of HbSS and/or its sequellae. (3) Test the hypothesis that erythroid cation channel dysrequlation of sickle cell disease is mirrored by similar dysregulation in red cells of mouse models of sickle cell disease. The experiments of this aim will extend the findings of Aim 1 to characterize the cation channel activity of mouse erythrocytes expressing with HbSAD in he presence of some mouse Hb or expressing almost entirely human HbS.